A high-efficiency CRISPR/Cas9 system for targeted mutagenesis in Cotton (Gossypium hirsutum L.)

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dc.contributor.authorZhang, Baohong
dc.contributor.authorLi, Chao
dc.contributor.authorUnver, Turgay
dc.date.accessioned2017-08-23T15:10:06Z
dc.date.available2017-08-23T15:10:06Z
dc.date.issued2017-03-03
dc.description.abstractThe complex allotetraploid genome is one of major challenges in cotton for repressing gene expression. Developing site-specific DNA mutation is the long-term dream for cotton breeding scientists. The clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) system is emerging as a robust biotechnology for targeted-DNA mutation. In this study, two sgRNAs, GhMYB25-like-sgRNA1 and GhMYB25-like-sgRNA2, were designed in the identical genomic regions of GhMYB25-like A and GhMYB25-like D, which were encoded by cotton A subgenome and the D subgenome, respectively, was assembled to direct Cas9-mediated allotetraploid cotton genome editing. High proportion (14.2–21.4%) CRISPR/Cas9-induced specific truncation events, either from GhMYB25-like A DNA site or from GhMYB25-like D DNA site, were detected in 50% examined transgenic cotton through PCR amplification assay and sequencing analyses. Sequencing results also demonstrated that 100% and 98.8% mutation frequency were occurred on GhMYB25-like-sgRNA1 and GhMYB25-like-sgRNA2 target site respectively. The off-target effect was evaluated by sequencing two putative off-target sites, which have 3 and 1 mismatched nucleotides with GhMYB25-like-sgRNA1 and GhMYB25-like-sgRNA2, respectively; all the examined samples were not detected any off-targetcaused mutation events. Thus, these results demonstrated that CRISPR/Cas9 is qualified for generating DNA level mutations on allotetraploid cotton genome with high-efficiency and high-specificity.en_US
dc.description.sponsorshipECU Open Access Publishing Support Funden_US
dc.identifier.citationLi, C. et al. A high-efficiency CRISPR/Cas9 system for targeted mutagenesis in Cotton (Gossypium hirsutum L.). Sci. Rep. 7, 43902; doi: 10.1038/srep43902 (2017).en_US
dc.identifier.doi10.1038/srep43902
dc.identifier.urihttp://hdl.handle.net/10342/6405
dc.language.isoen_USen_US
dc.relation.urihttps://doi.org/10.1038/srep43902en_US
dc.titleA high-efficiency CRISPR/Cas9 system for targeted mutagenesis in Cotton (Gossypium hirsutum L.)en_US
dc.typeArticleen_US
ecu.journal.issue43902en_US
ecu.journal.nameScientific Reportsen_US
ecu.journal.volume7en_US

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